Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase (RTK) in the insulin receptor superfamily initially identified in one of its constitutively activated oncogenic fusion forms—nucleophosmin (NPM)-ALK—in anaplastic large-cell lymphomas (Morris et al. (1994) Science 263:1281-1284; Morris et al. (1997) Oncogene 14:2175-2188). Subsequent studies have identified ALK fusions in subsets of diffuse large B-cell lymphomas, malignant histiocytosis, inflammatory myofibroblastic tumor sarcomas, esophageal squamous cell carcinomas, breast cancers, colorectal carcinomas, and non-small cell lung carcinomas (reviewed in Webb et al. (2009) Expert Rev Anticancer Ther 9:331-356). Most recently, genomic DNA amplification and protein overexpression, as well as activating point mutations, of ALK have been shown to cause neuroblastomas (Webb et al. (2009) Expert Rev Anticancer Ther 9:331-356; George et al. (2008) Nature 455:975-979). In addition to these cancers for which a causative role for aberrant ALK activity is well validated, more circumstantial links implicate ALK in the genesis of yet other malignancies, such as glioblastoma, via a mechanism of receptor activation involving autocrine and/or paracrine growth loops with the reported ALK ligands, pleiotrophin and midkine (Webb et al. (2009) Expert Rev Anticancer Ther 9:331-356).
The involvement of mutant, constitutively activated forms of ALK in this broad spectrum of cancers has engendered considerable interest among pharmaceutical and biotech firms in the development of ALK inhibitors analogous to the small-molecule kinase inhibitors imatinib (Gleevec, Novartis) and erlotinib (Tarceva, Genentech/OSI) that target the Abelson (ABL) kinase and the epidermal growth factor receptor (EGFR) kinase, respectively. Since 2001, eight ATP-competitive small-molecule kinase inhibitors (including imatinib and erlotinib) have been approved for various cancer indications in the United States (reviewed in Webb et al. (2009) Expert Rev Anticancer Ther 9:331-356). Although these drugs have proven extremely valuable as anticancer agents—perhaps best exemplified by the therapeutic benefit realized in patients with chronic myeloid leukemia (CML) and gastrointestinal stromal tumors (GIST) following administration of imatinib mesylate (Gleevec, Novartis), the use of these inhibitors in the clinic has led to the emergence of drug-resistant tumors (O'Hare et al. (2007) Blood 110:2242-2249; Engelman and Settleman (2008) Curr Opin Genet Dev 18:1-7; Bikker et al. (2009) J Med Chem 52:1493-1509). This resistance has been attributed to a number of mechanisms including the amplification of the gene encoding the oncogenic kinase as well as the activation of alternative signaling pathways; however, the most common mechanism mediating ATP-competitive kinase inhibitor resistance is the development of individual or groups of mutations within or near the kinase catalytic domains of the kinase targets (O'Hare et al. (2007) Blood 110:2242-2249; Engelman and Settleman (2008) Curr Opin Genet Dev 18:1-7; Bikker et al. (2009) J Med Chem 52:1493-1509). These mutations preclude high-affinity interactions of the inhibitors with their kinase targets while leaving ATP binding by their catalytic domains intact. The emergence of clinical resistance to kinase inhibitors and identification of the kinase domain mutations that confer such resistance have engendered the design and development of follow-on drugs to treat patients whose tumors no longer respond to therapy with first-generation agents
Robust diagnostic assays to detect the presence of resistance mutations in the ALK kinase domain are needed for clinical application to confirm the mechanism of resistance in cancer patients who become resistant to therapy with ALK kinase inhibitors, and to permit the informed selection by physicians of second-generation inhibitors for the management of patients with first-generation inhibitor-resistant tumors. No assays for the detection of ALK inhibitor resistance currently exist. The identification of these mutations will also serve to guide the informed design and synthesis of second- and later-generation inhibitors of ALK developed to inhibit these mutant forms of ALK that are resistant to first-generation inhibitors.